title: Structural basis for the selective inhibition of human 3beta-hydroxysteroid dehydrogenase 1 in human breast tumor MCF-7 cells.
abstract: Human 3beta-hydroxysteroid dehydrogenase/isomerase type 1 (3beta-HSD1) is a critical enzyme in the conversion of DHEA to estradiol in breast tumors and may be a target enzyme for inhibition in the treatment of breast cancer in postmenopausal women. Human 3beta-HSD2 participates in the production of cortisol and aldosterone in the human adrenal gland in this population. In our recombinant human breast tumor MCF-7 Tet-off cells that express either 3beta-HSD1 or 3beta-HSD2, trilostane and epostane inhibit the DHEA-induced proliferation of MCF-7 3beta-HSD1 cells with 12- to 16-fold lower IC(50) values compared to the MCF-7 3beta-HSD2 cells. The compounds also competitively inhibit purified human 3beta-HSD1 with 12- to 16-fold lower K(i) values compared to the noncompetitive K(i) values measured for human 3beta-HSD2. Using our structural model of 3beta-HSD1, trilostane or 17beta-acetoxy-trilostane was docked in the active site of 3beta-HSD1, and Arg195 in 3beta-HSD1 or Pro195 in 3beta-HSD2 was identified as a potentially critical residue (one of 23 non-identical residues in the two isoenzymes). The P195R mutant of 3beta-HSD2 were created, expressed and purified. Kinetic analyses of enzyme inhibition suggest that the high affinity, competitive inhibition of 3beta-HSD1 by trilostane and epostane may be related to the presence of Arg195 in 3beta-HSD1 vs. Pro195 in 3beta-HSD2.
authors.affiliation: Division of Basic Medical Sciences, Mercer University School of Medicine, Mercer University, 1550 College Street, Macon, GA 31207, USA. thomas.j@Mercer.edu
authors.forename: James L
authors.initials: JL
authors.lastname: Thomas
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Bucholtz
authors.forename: Jingping
authors.initials: J
authors.lastname: Sun
authors.forename: Vance L
authors.initials: VL
authors.lastname: Mack
authors.forename: Balint
authors.initials: B
authors.lastname: Kacsoh
mesh_terms: D015096:3-Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D001120:Arginine / Q000378:metabolism; D047072:Aromatase Inhibitors / Q000494:pharmacology; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000201:enzymology* / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003067:Coenzymes / Q000378:metabolism; D003687:Dehydroepiandrosterone / Q000494:pharmacology; D013196:Dihydrotestosterone / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004958:Estradiol / Q000494:pharmacology; D004970:Estrone / Q000494:pharmacology; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D050505:Mutant Proteins / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013379:Substrate Specificity / Q000187:drug effects
chemical_list: D047072:Aromatase Inhibitors; D003067:Coenzymes; D004791:Enzyme Inhibitors; D050505:Mutant Proteins; D012333:RNA, Messenger; D013196:Dihydrotestosterone; D004970:Estrone; D003687:Dehydroepiandrosterone; D004958:Estradiol; D001120:Arginine; D015096:3-Hydroxysteroid Dehydrogenases; C009954:trilostane